Study on effect of echinococcus granulosus protoscolices on fibrosis of bone marrow mesenchymal stem cells_Chinese Journal of Reparative and Reconstructive Surgery

Authors：

WU Jie ¹ , GUI Xianwei ² , JIANG Huijiao ² , LIANG Xueqi ² , WANG Erqiang ² , XU Xiaodan ² , CHEN Xueling ² ,  WU Xiangwei ¹

1. Department of Hepatobiliary Surgery, the First Affiliated Hospital, School of Medicine, Shihezi University, Shihezi Xinjiang, 832008, P.R.China;
2. Department of Immunology, School of Medicine, Shihezi University, Shihezi Xinjiang, 832008, P.R.China;

Corresponding author：

WU Xiangwei, Email: wxwshz@126.com

Keywords：

Bone marrow mesenchymal stem cells; fibrosis; echinococcus granulosus; transforming growth factor β₁; collagen type Ⅰ; collagen type Ⅲ; signaling pathway

DOI：

10.7507/1002-1892.201909050

Video：

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Objective To investigate the effect of echinococcus granulosus protoscolices on the differentiation of bone marrow mesenchymal stem cells (BMSCs) into fibroblasts.Methods Femur bone marrow of 4-week-old C57BL/6 mice was taken and BMSCs were isolated and cultured by adherent culture. Echinococcus granulosus protoscolices was extracted from the liver of sheep infected with echinococcus granulosus. The experiment was divided into two groups. The experimental group was co-cultured with the 3rd generation BMSCs and the echinococcus granulosus protoscolices, and the control group was the 3rd generation BMSCs. Before and after co-culture, the morphology of BMSCs and the activity of echinococcus granulosus protoscolices were observed by inverted microscope. After cultured for 1, 3, 5, and 7 days, the mRNA expressions of transforming growth factor β₁ (TGF-β₁), collagen type Ⅰ, and collagen type Ⅲ were detected by real-time fluorescent quantitative PCR, the protein expressions of TGF-β₁, collagen type Ⅰ, collagen type Ⅲ, Smad7, and phosphorylated Smad2/3 were detected by Western blot, and the contents of collagen type Ⅰ and collagen type Ⅲ in the supernatant of the two groups were detected by ELISA.Results After 7 days of co-culture, the morphology of BMSCs changed into fusiform and irregular triangle, which was closer to the mouse fibroblasts. The relative mRNA expressions of TGF-β₁, collagen type Ⅰ, and collagen type Ⅲ in the experimental group were significantly higher than those in the control group; the relative protein expressions of TGF-β₁, collagen type Ⅰ, collagen type Ⅲ, and phosphorylated Smad2/3 in the experimental group were significantly higher than those in the control group, and the relative protein expression of Smad7 in the experimental group was significantly lower than that in the control group; the contents of collagen type Ⅰ and collagen type Ⅲ in the supernatant of the experimental group were significantly higher than those in the control group. The differences between the two groups were significant (P<0.05).Conclusion Echinococcus granulosus protoscolices may promote the secretion of collagen type Ⅰ, collagen type Ⅲ, and TGF-β₁ by TGF-β₁/Smad signal pathway, which can promote the fibrosis of BMSCs that related to the formation of fibrocystic wall by echinococcosis.

Citation： WU Jie, GUI Xianwei, JIANG Huijiao, LIANG Xueqi, WANG Erqiang, XU Xiaodan, CHEN Xueling, WU Xiangwei. Study on effect of echinococcus granulosus protoscolices on fibrosis of bone marrow mesenchymal stem cells. Chinese Journal of Reparative and Reconstructive Surgery, 2020, 34(5): 630-636. doi: 10.7507/1002-1892.201909050 Copy

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2.	陈小林, 印双红, 吴向未, 等. 原头蚴及囊液促体外培养小鼠脾细胞 TGF-β 表达的研究. 中国病原生物学杂志, 2013, 8(6): 523-526.
3.	Coque TM, Willems R, Cantón R, et al. High occurrence of esp among ampicillin-resistant and vancomycin-susceptible Enterococcus faecium clones from hospitalized patients. J Antimicrob Chemother, 2002, 50(6): 1035-1038.
4.	Czermak BV, Akhan O, Hiemetzberger R, et al. Echinococcosis of the liver. Abdom Imaging, 2008, 33(2): 133-143.
5.	Herzog EL, Chai L, Krause DS. Plasticity of marrow-derived stem cells. Blood, 2003, 102(10): 3483-3493.
6.	王福科, 张红, 李彦林, 等. 联合培养血管内皮细胞与脂肪干细胞构建组织工程骨异位成骨. 中国修复重建外科杂志, 2019, 33(10): 1310-1319.
7.	Bernardo ME, Emons JA, Karperien M, et al. Human mesenchymal stem cells derived from bone marrow display a better chondrogenic differentiation compared with other sources. Connect Tissue Res, 2007, 48(3): 132-140.
8.	Lee HY, Hong IS. Double-edged sword of mesenchymal stem cells: Cancer-promoting versus therapeutic potential. Cancer Sci, 2017, 108(10): 1939-1946.
9.	Ma S, Xie N, Li W, et al. Immunobiology of mesenchymal stem cells. Cell Death Differ, 2014, 21(2): 216-225.
10.	廖筱梅, 罗兴前, 代蕾, 等. 脂肪间充质干细胞-壳聚糖凝胶复合物治疗大鼠深 Ⅱ 度烫伤创面实验研究. 中国修复重建外科杂志, 2019, 33(1): 101-109.
11.	Kim KK, Sheppard D, Chapman HA. TGF-β1 signaling and tissue fibrosis. Cold Spring Harb Perspect Biol, 2018, 10(4): pii: a022293.
12.	Shi Y, Massagué J. Mechanisms of TGF-beta signaling from cell membrane to the nucleus. Cell, 2003, 113(6): 685-700.
13.	Wang X, Zhao Z, Zhang H, et al. Simultaneous isolation of mesenchymal stem cells and endothelial progenitor cells derived from murine bone marrow. Exp Ther Med, 2018, 16(6): 5171-5177.
14.	胡素贤, 倪晓妍, 王嘉逊, 等. 新生小鼠皮肤成纤维细胞的分离培养与鉴定. 中国医药导报, 2011, 8(29): 5-7, 26, 封 3.
15.	Yin CM, Suen WC, Lin S, et al. Dysregulation of both miR-140-3p and miR-140-5p in synovial fluid correlate with osteoarthritis severity. Bone Joint Res, 2017, 6(11): 612-618.
16.	Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods, 2001, 25(4): 402-408.
17.	Vuitton DA. The ambiguous role of immunity in echinococcosis: protection of the host or of the parasite? Acta Trop, 2003, 85(2): 119-132.
18.	Tang Y, Wu X, Lei W, et al. TGF-beta1-induced migration of bone mesenchymal stem cells couples bone resorption with formation. Nat Med, 2009, 15(7): 757-765.
19.	Ask K, Bonniaud P, Maass K, et al. Progressive pulmonary fibrosis is mediated by TGF-beta isoform 1 but not TGF-beta3. Int J Biochem Cell Biol, 2008, 40(3): 484-495.
20.	Johnston EF, Gillis TE. Transforming growth factor beta-1 (TGF-β1) stimulates collagen synthesis in cultured rainbow trout cardiac fibroblasts. J Exp Biol, 2017, 220(Pt 14): 2645-2653.
21.	Kim YJ, Yoo SM, Park HH, et al. Exosomes derived from human umbilical cord blood mesenchymal stem cells stimulates rejuvenation of human skin. Biochem Biophys Res Commun, 2017, 493(2): 1102-1108.
22.	张旭勇, 夏杰, 杨涛, 等. TGFβR Ⅰ 和 TGFβR Ⅱ 在肝细粒棘球蚴外囊壁中的表达. 医学研究生学报, 2016, 29(1): 66-69.
23.	印双红, 陈小林, 徐芳洁, 等. 小鼠感染细粒棘球蚴早期 TGF-β1 表达分析. 中国病原生物学杂志, 2014, 9(2): 155-157.
24.	印双红, 张俊波, 陈小林, 等. 细粒棘球蚴感染中阻断 TGF-β1 受体对淋巴细胞的影响. 中国免疫学杂志, 2015, 31(5): 607-612.
25.	Deplazes P, Rinaldi L, Alvarez Rojas CA, et al. Global distribution of alveolar and cystic echinococcosis. Adv Parasitol, 2017, 95: 315-493.

1. Li T, Ito A, Nakaya K, et al. Species identification of human echinococcosis using histopathology and genotyping in northwestern China. Trans R Soc Trop Med Hyg, 2008, 102(6): 585-590.
2. 陈小林, 印双红, 吴向未, 等. 原头蚴及囊液促体外培养小鼠脾细胞 TGF-β 表达的研究. 中国病原生物学杂志, 2013, 8(6): 523-526.
3. Coque TM, Willems R, Cantón R, et al. High occurrence of esp among ampicillin-resistant and vancomycin-susceptible Enterococcus faecium clones from hospitalized patients. J Antimicrob Chemother, 2002, 50(6): 1035-1038.
4. Czermak BV, Akhan O, Hiemetzberger R, et al. Echinococcosis of the liver. Abdom Imaging, 2008, 33(2): 133-143.
5. Herzog EL, Chai L, Krause DS. Plasticity of marrow-derived stem cells. Blood, 2003, 102(10): 3483-3493.
6. 王福科, 张红, 李彦林, 等. 联合培养血管内皮细胞与脂肪干细胞构建组织工程骨异位成骨. 中国修复重建外科杂志, 2019, 33(10): 1310-1319.
7. Bernardo ME, Emons JA, Karperien M, et al. Human mesenchymal stem cells derived from bone marrow display a better chondrogenic differentiation compared with other sources. Connect Tissue Res, 2007, 48(3): 132-140.
8. Lee HY, Hong IS. Double-edged sword of mesenchymal stem cells: Cancer-promoting versus therapeutic potential. Cancer Sci, 2017, 108(10): 1939-1946.
9. Ma S, Xie N, Li W, et al. Immunobiology of mesenchymal stem cells. Cell Death Differ, 2014, 21(2): 216-225.
10. 廖筱梅, 罗兴前, 代蕾, 等. 脂肪间充质干细胞-壳聚糖凝胶复合物治疗大鼠深 Ⅱ 度烫伤创面实验研究. 中国修复重建外科杂志, 2019, 33(1): 101-109.
11. Kim KK, Sheppard D, Chapman HA. TGF-β1 signaling and tissue fibrosis. Cold Spring Harb Perspect Biol, 2018, 10(4): pii: a022293.
12. Shi Y, Massagué J. Mechanisms of TGF-beta signaling from cell membrane to the nucleus. Cell, 2003, 113(6): 685-700.
13. Wang X, Zhao Z, Zhang H, et al. Simultaneous isolation of mesenchymal stem cells and endothelial progenitor cells derived from murine bone marrow. Exp Ther Med, 2018, 16(6): 5171-5177.
14. 胡素贤, 倪晓妍, 王嘉逊, 等. 新生小鼠皮肤成纤维细胞的分离培养与鉴定. 中国医药导报, 2011, 8(29): 5-7, 26, 封 3.
15. Yin CM, Suen WC, Lin S, et al. Dysregulation of both miR-140-3p and miR-140-5p in synovial fluid correlate with osteoarthritis severity. Bone Joint Res, 2017, 6(11): 612-618.
16. Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods, 2001, 25(4): 402-408.
17. Vuitton DA. The ambiguous role of immunity in echinococcosis: protection of the host or of the parasite? Acta Trop, 2003, 85(2): 119-132.
18. Tang Y, Wu X, Lei W, et al. TGF-beta1-induced migration of bone mesenchymal stem cells couples bone resorption with formation. Nat Med, 2009, 15(7): 757-765.
19. Ask K, Bonniaud P, Maass K, et al. Progressive pulmonary fibrosis is mediated by TGF-beta isoform 1 but not TGF-beta3. Int J Biochem Cell Biol, 2008, 40(3): 484-495.
20. Johnston EF, Gillis TE. Transforming growth factor beta-1 (TGF-β1) stimulates collagen synthesis in cultured rainbow trout cardiac fibroblasts. J Exp Biol, 2017, 220(Pt 14): 2645-2653.
21. Kim YJ, Yoo SM, Park HH, et al. Exosomes derived from human umbilical cord blood mesenchymal stem cells stimulates rejuvenation of human skin. Biochem Biophys Res Commun, 2017, 493(2): 1102-1108.
22. 张旭勇, 夏杰, 杨涛, 等. TGFβR Ⅰ 和 TGFβR Ⅱ 在肝细粒棘球蚴外囊壁中的表达. 医学研究生学报, 2016, 29(1): 66-69.
23. 印双红, 陈小林, 徐芳洁, 等. 小鼠感染细粒棘球蚴早期 TGF-β1 表达分析. 中国病原生物学杂志, 2014, 9(2): 155-157.
24. 印双红, 张俊波, 陈小林, 等. 细粒棘球蚴感染中阻断 TGF-β1 受体对淋巴细胞的影响. 中国免疫学杂志, 2015, 31(5): 607-612.
25. Deplazes P, Rinaldi L, Alvarez Rojas CA, et al. Global distribution of alveolar and cystic echinococcosis. Adv Parasitol, 2017, 95: 315-493.

Chinese Journal of Reparative and Reconstructive Surgery

Study on effect of echinococcus granulosus protoscolices on fibrosis of bone marrow mesenchymal stem cells

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